Protein/Peptide Transduction in Metanephric Explant Culture

Abstract

While gene targeting methods have largely supplanted cell/explant culture models for studying developmental processes, they have not eliminated the need for or value of such approaches in the investigator’s technical arsenal. Explant culture models, such as those devised for the metanephric kidney and its progenitors, remain invaluable as tools for screening regulatory factors involved in tissue induction or in the inhibition of progenitor specification. Thus, some factors capable of inducing tissue condensations or nephronic tubule formation in explants of metanephric mesenchyme have been identified through direct treatment of cultures rather than lengthy genetic engineering in animals. Unfortunately, renal progenitors are largely refractory to most contemporary methods for gene manipulation, including transfection and viral transduction, so the applications of explant culture have been rather limited. However, methods for protein or peptide transduction offer greatly improved efficiencies for uptake and expression/regulation of proteins within cells and tissues. Biologically active TAT- or penetratin-fusion proteins/peptides are readily taken up by most cells in metanephric explants or monolayer cultured cells (Plisov et al., J Am Soc Nephrol 16:1632–1644, 2005; Osafune et al., Development 133:151–161, 2006; Wang et al., Cell Signal 22:1717–1726, 2010; Tanigawa, Dev Biol 352:58–69, 2011), allowing a direct functional evaluation of theoretically any protein, including biologically active enzymes and transcription factors, or any targeted interactive domain within a protein.